The long-range objective of the proposed research is to determine the cellular and molecular events that lead to the differentiation of specific cell types in the vertebrate retina. The primary goal of the current application is to evaluate a mechanistic model for how Hedgehog (Hh) signaling and retinoic acid (RA) signaling control photoreceptor development. In this model, these signaling systems are co-regulated and act in parallel: Hh signaling propagates cone differentiation and regulates RA signaling, and RA signaling propagates rod differentiation and regulates photoreceptor phenotype. We hypothesize that Hh and RA act by influencing expression of the transcription factors crx and rxl/2, which in turn regulate photoreceptor-specific gene expression. Our research uses the zebrafish, a powerful, in vivo system for studying the mechanisms of vertebrate photoreceptor development. Through the use of genetic and molecular tools that allow the manipulation of the Hh and RA signaling systems, we will (1) identify cellular and molecular targets of Hh signaling during photoreceptor differentiation, (2) identify cellular and molecular targets of RA signaling during photoreceptor differentiation, and (3) determine the mechanism by which Hh and RA signaling are coordinated. In addition to testing these components of the mechanistic model, our experiments will determine whether Hh and RA influence photoreceptor cell fate. The mechanisms that generate multiple neuronal cell types in the correct ratios present important and challenging issues in neurobiology. The proposed research is expected to elucidate the respective roles of Hh and RA in regulating development of specific photoreceptor types in vivo. Furthermore, retinoid treatment is under active testing as a therapy for retinal disease, and Hh has been suggested as a potential treatment for specific visual disorders. An improved understanding of the mechanisms of action of Hh and RA will be valuable for reassessing current treatments, and for designing future therapies.